Icariin accelerates bone regeneration by inducing osteogenesisangiogenesis coupling in rats with type 1 diabetes mellitus

BACKGROUND Icariin(ICA),a natural flavonoid compound monomer,has multiple pharmacological activities.However,its effect on bone defect in the context of type 1 diabetes mellitus(T1DM)has not yet been examined.AIM To explore the role and potential mechanism of ICA on bone defect in the context of T1DM.METHODS The effects of ICA on osteogenesis and angiogenesis were evaluated by alkaline phosphatase staining,alizarin red S staining,quantitative real-time polymerase chain reaction,Western blot,and immunofluorescence.Angiogenesis-related assays were conducted to investigate the relationship between osteogenesis and angiogenesis.A bone defect model was established in T1DM rats.The model rats were then treated with ICA or placebo and micron-scale computed tomography,histomorphometry,histology,and sequential fluorescent labeling were used to evaluate the effect of ICA on bone formation in the defect area.RESULTS ICA promoted bone marrow mesenchymal stem cell(BMSC)proliferation and osteogenic differentiation.The ICA treated-BMSCs showed higher expression levels of osteogenesis-related markers(alkaline phosphatase and osteocalcin)and angiogenesis-related markers(vascular endothelial growth factor A and platelet endothelial cell adhesion molecule 1)compared to the untreated group.ICA was also found to induce osteogenesis-angiogenesis coupling of BMSCs.In the bone defect model T1DM rats,ICA facilitated bone formation and CD31hiEMCNhi type H-positive capillary formation.Lastly,ICA effectively accelerated the rate of bone formation in the defect area.CONCLUSION ICA was able to accelerate bone regeneration in a T1DM rat model by inducing osteogenesis-angiogenesis coupling of BMSCs.

Icariin plus curcumol enhances autophagy through the mTOR pathway and promotes cathepsin B-mediated pyroptosis of prostate cancer cells

Objective:To examine the effect of icariin plus curcumol on prostate cancer cells PC3 and elucidate the underlying mechanisms.Methods:We employed the Cell Counting Kit 8 assay and colony formation assay to assess cell viability and proliferation.Autophagy expression was analyzed using monodansylcadaverine staining.Immunofluorescence and Western blot analyses were used to evaluate protein expressions related to autophagy,pyroptosis,and the mTOR pathway.Cellular damage was examined using the lactate dehydrogenase assay.Moreover,cathepsin B and NLRP3 were detected by co-immunoprecipitation.Results:Icariin plus curcumol led to a decrease in PC3 cell proliferation and an enhancement of autophagy.The levels of LC3-Ⅱ/LC3-Ⅰand beclin-1 were increased,while the levels of p62 and mTOR were decreased after treatment with icariin plus curcumol.These changes were reversed upon overexpression of mTOR.Furthermore,3-methyladenine resulted in a decrease in inflammatory cytokines,pyroptosis-related protein levels,and lactate dehydrogenase concentration,compared to the icariin plus curcumol group.Inhibiting cathepsin B reversed the regulatory effects of icariin plus curcumol.Conclusions:Icariin plus curcumol demonstrates great potential as a therapeutic agent for castration-resistant prostate cancer by enhancing autophagy via the mTOR pathway and promoting pyroptosis mediated by cathepsin B.These findings provide valuable insights into the molecular mechanisms underlying the therapeutic potential of icariin and curcumol for prostate cancer treatment.

Icariin ameliorates viral myocarditis by inhibiting TLR4-mediated ferroptosis

Objective:To explore the mechanism by which icariin alleviates viral myocarditis.Methods:CVB3-induced cardiomyocytes were used as an in vitro model of viral myocarditis to assess the effects of icariin treatment on cell viability,inflammation,and apoptosis.Moreover,the effects of icariin on ferroptosis and TLR4 signaling were assessed.After AC16 cells were transfected with TLR4 overexpression plasmids,the role of TLR4 in mediating the regulatory effect of icariin in viral myocarditis was investigated.Results:Icariin significantly elevated cell viability and reduced inflammatory factors TNF-α,IL-1β,IL-6,and IL-18.Flow cytometry revealed that icariin decreased apoptosis rate,and the protein expression of Bax and cleaved caspase 3 and 9 in CVB3-induced cardiomyocytes.Additionally,it suppressed ferroptosis including lipid peroxidation and ferrous ion,as well as the TLR4 signaling.However,TLR4 overexpression abrogated the modulatory effects of icariin.Conclusions:Icariin mitigates CVB3-induced myocardial injury by inhibiting TLR4-mediated ferroptosis.Further animal study is needed to verify its efficacy.

Icariin Ameliorates D-galactose-induced Cell Injury in Neuron-like PC12 Cells by Inhibiting MPTP Opening

Objective Icariin(ICA)has a good neuroprotective effect and can upregulate neuronal basal autophagy in naturally aging rats.Mitochondrial dysfunction is associated with brain aging-related neurodegenerative diseases.Abnormal opening of the mitochondrial permeability transition pore(mPTP)is a crucial factor in mitochondrial dysfunction and is associated with excessive autophagy.This study aimed to explore that ICA protects against neuronal injury by blocking the mPTP opening and down-regulating autophagy levels in a D-galactose(D-gal)-induced cell injury model.Methods A cell model of neuronal injury was established in rat pheochromocytoma cells(PC12 cells)treated with 200 mmol/L D-gal for 48 h.In this cell model,PC12 cells were pre-treated with different concentrations of ICA for 24 h.MTT was used to detect cell viability.Senescence associatedβ-galactosidase(SA-β-Gal)staining was used to observe cell senescence.Western blot analysis was performed to detect the expression levels of a senescence-related protein(p21),autophagy markers(LC3B,p62,Atg7,Atg5 and Beclin 1),mitochondrial fission and fusion-related proteins(Drp1,Mfn2 and Opa1),and mitophagy markers(Pink1 and Parkin).The changes of autophagic flow were detected by using mRFP-GFP-LC3 adenovirus.The intracellular ultrastructure was observed by transmission electron microscopy.Immunofluorescence was used to detect mPTP,mitochondrial membrane potential(MMP),mitochondrial reactive oxygen species(mtROS)and ROS levels.ROS and apoptosis levels were detected by flow cytometry.Results D-gal treatment significantly decreased the viability of PC12 cells,and markedly increased the SA-β-Gal positive cells as compared to the control group.With the D-gal stimulation,the expression of p21 was significantly up-regulated.Furthermore,D-gal stimulation resulted in an elevated LC3B II/I ratio and decreased p62 expression.Meanwhile,autophagosomes and autolysosomes were significantly increased,indicating abnormal activation of autophagy levels.In addition,in this D-gal-induced model of cell injury,the mPTP was abnormally open,the ROS generation was continuously increased,the MMP was gradually decreased,and the apoptosis was increased.ICA effectively improved mitochondrial dysfunction to protect against D-gal-induced cell injury and apoptosis.It strongly inhibited excessive autophagy by blocking the opening of the mPTP.Cotreatment with ICA and an mPTP inhibitor(cyclosporin A)did not ameliorate mitochondrial dysfunction.However,the protective effects were attenuated by cotreatment with ICA and an mPTP activator(lonidamine).Conclusion ICA inhibits the activation of excessive autophagy and thus improves mitochondrial dysfunction by blocking the mPTP opening.

Bibliometric analysis of 100 most-cited papers on Icariin

Icariin is the most prevalent component of the medicinal herb Herba Epimedii.Icariin exhibits many medicinal properties,including anti-cancer impact and osteoprotective and neuroprotective effects.The goal of this study was to use bibliometric analysis to find and describe the top 100 papers about Icariin that had received the most citations.The Science Citation Index-Expanded(SCI-E)of the Web of Science Core Collection was used to find publications on Icariin(WoSCC).Descriptive analysis was conducted using VOSviewer software.There were 1473 articles about Icariin in all.The top 100 papers were published between 1996 and 2024 and received citations in the range of 55 to 390.The country that has contributed the most to Icariin research is China(84).The most productive institution was Fudan University.The most published journal was Phytomedicine.The research hotspots of Icariin mainly focus on the following aspects:research on Icariin treatment of sex hormone-related osteoporosis and erectile function;The effect of Icariin on cells by regulating oxidative stress,apoptosis,and proliferation;the mechanism of Icariin in the treatment of cancer;the neuroprotective effect of Icariin in central nervous diseases,such as Alzheimer's disease,Parkinson's disease,and depression.Future research should focus on further elucidating Icariin's anti-tumor effects,its application in cartilage tissue engineering and orthopedic biomaterials,and developing novel drug delivery systems to enhance its bioavailability.This research contributed essential knowledge to the study of Icariin.These results may be used in new study areas and to direct drug development.

Analysis of the Antidepressant Effects and Mechanisms of Icariin II Based on The GABAergic Nervous System

This article summarized the research progress on the antidepressant mechanism of icariin II,mainly elaborating on its mechanism from five aspects:GABAergic nervous system,inflammatory response,oxidative stress,neurotrophic factors,and neurotransmitters in the brain.Its clinical application value was further explored to provide a theoretical basis for the development and utilization of icariin II in treating depression.

Bone mesenchymal stem cell-derived exosomes involved co-delivery and synergism effect with icariin via mussel-inspired multifunctional hydrogel for cartilage protection

Mesenchymal stem cells(MSC)are particularly effective in promoting cartilage regeneration due to their immunomodulatory,anti-inflammatory and regenerative repair functions of tissues and organs.Meanwhile,the intra-articular delivery and synergy with other therapeutic drugs have been the key issues driving their further application.We report a mussel-inspired multifunctional hydrogel system,which could achieve co-delivery and synergism effect of MSC-derived exosomes(Exos)with icariin(ICA).The ICA and Exos co-delivered articular cavity injection system are expected to retain in the joint cavity and promote cartilage regeneration,due to the thermosensitive,self-healing and adhesion properties of the mussel-inspired multifunctional hydrogel.The experimental results proved that Exos enhanced the cellular uptake of ICA by more than 2-fold evenly,and the synergism of Exos and ICA efficiently improve the cell proliferation and migration.After synergic treatment,the content of matrix metalloproteinase 13 in the supernatant and intracellular decreased by 47%and 59%,respectively.In vivo study,ICA-loaded Exos exhibited prolonged retention behavior bymultifunctional hydrogel delivery,thus displayed an increased cartilage protection.In the model of osteoarthritis,co-delivery hydrogel system relieved the cartilage recession,ensuring appropriate cartilage thickness.

Icariin ameliorates memory deficits through regulating brain insulin signaling and glucose transporters in 3×Tg-AD mice

Icariin,a major prenylated flavonoid found in Epimedium spp.,is a bioactive constituent of Herba Epimedii and has been shown to exert neuroprotective effects in experimental models of Alzheimer’s disease.In this study,we investigated the neuroprotective mechanism of icariin in an APP/PS1/Tau triple-transgenic mouse model of Alzheimer’s disease.We performed behavioral tests,pathological examination,and western blot assay,and found that memory deficits of the model mice were obviously improved,neuronal and synaptic damage in the cerebral cortex was substantially mitigated,and amyloid-βaccumulation and tau hyperphosphorylation were considerably reduced after 5 months of intragastric administration of icariin at a dose of 60 mg/kg body weight per day.Furthermore,deficits of proteins in the insulin signaling pathway and their phosphorylation levels were significantly reversed,including the insulin receptor,insulin receptor substrate 1,phosphatidylinositol-3-kinase,protein kinase B,and glycogen synthase kinase 3β,and the levels of glucose transporter 1 and 3 were markedly increased.These findings suggest that icariin can improve learning and memory impairments in the mouse model of Alzheimer’s disease by regulating brain insulin signaling and glucose transporters,which lays the foundation for potential clinical application of icariin in the prevention and treatment of Alzheimer’s disease.

Potential role of microRNA-503 in Icariin-mediated prevention of high glucose-induced endoplasmic reticulum stress

BACKGROUND Dysregulated microRNA(miRNA)is crucial in the progression of diabetic nephropathy(DN).AIM To investigate the potential molecular mechanism of Icariin(ICA)in regulating endoplasmic reticulum(ER)stress-mediated apoptosis in high glucose(HG)-induced primary rat kidney cells(PRKs),with emphasis on the role of miR-503 and sirtuin 4(SIRT4)in this process.METHODS Single intraperitoneal injection of streptozotocin(65 mg/kg)in Sprague-Dawley rats induce DN in the in vivo hyperglycemic model.Glucose-treated PRKs were used as an in vitro HG model.An immunofluorescence assay identified isolated PRKs.Cell Counting Kit-8 and flow cytometry analyzed the effect of ICA treatment on cell viability and apoptosis,respectively.Real-time quantitative polymerase chain reaction and western blot analyzed the levels of ER stressrelated proteins.Dual luciferase analysis of miR-503 binding to downstream SIRT4 was performed.RESULTS ICA treatment alleviated the upregulated miR-503 expression in vivo(DN)and in vitro(HG).Mechanistically,ICA reduced HG-induced miR-503 overexpression,thereby counteracting its function in downregulating SIRT4 levels.ICA regulated the miR-503/SIRT4 axis and subsequent ER stress to alleviate HG-induced PRKs injury.CONCLUSION ICA reduced HG-mediated inhibition of cell viability,promotion of apoptosis,and ER stress in PRKs.These effects involved regulation of the miR-503/SIRT4 axis.These findings indicate the potential of ICA to treat DN,and implicate miR-503 as a viable target for therapeutic interventions in DN.

Exploring the mechanism of icariin in regulat⁃ing cardiac microvascular endothelial cells based on network pharmacology,molecular docking and in vitro experiments

OBJECTIVE To investigate the regulatory effects of icariin(ICA)on cardiac micro⁃vascular endothelial cells(CMEC)after oxygenglucose deprivation reperfusion(OGD/R)injury.METHODS CMEC were subjected to OGD/R treatment to construct a myocardial ischemiareperfusion model,and were divided into normal,model,low(10μmol·L^(-1)),medium(20μmol·L^(-1))and high(40μmol·L^(-1))ICA group,and high ICA+inhibitor group(40μmol·L^(-1)+20 nmol·L^(-1)).CCK-8 assay was used to assess the protective ability of ICA against CMEC,and cell migration assay and tube-formation assay were used to detect the migration and generation ability of CMEC.The TCMSP database,Swiss-Target database and literature mining methods were used to col⁃lect ICA-related targets,the GeneCards data⁃base was used to collect target genes related to myocardial ischemia/reperfusion,and Cytoscape 3.8.0 software was used to construct a"drug-tar⁃get-disease"network.The potential targets were imported into STRING 11.5 database to obtain the PPI network.GO and KEGG enrichment analyses were performed on the potential targets using the DAVID database.Molecular docking was performed using AutoDock-vina 1.1.2 soft⁃ware.Western blot detected the expression of related proteins.RESULTS After CMEC was subjected to OGD/R treatment,ICA had a protec⁃tive effect at 10^(-1)60μmol·L^(-1);the results of the cell migration assay showed that each group of ICA could promote the migratory effect of CMEC(P<0.01,P<0.01);and the results of tube-for⁃mation assay showed that each group of ICA could significantly promote the generation of branches(P<0.01)and the capillary length exten⁃sion(P<0.05).Network pharmacology collected a total of 23 ICA action targets,1500 disease tar⁃gets and 12 key targets.GO function enrichment analysis found 85 results.KEGG pathway enrich⁃ment analysis found 53 results,involving AGERAGE signaling pathway,sphingolipid signaling pathway and VEGF signaling pathway.Molecu⁃lar docking results showed that ICA had better binding with core targets PRKCB,PRKCA and PTGS2.Western blot results showed that ICA could regulate the expression of PRKCB,PRKCA and PTGS2 proteins.The results of cell migra⁃tion assay,tube-formation assay and protein expression were reversed after addition of PKC inhibitor.CONCLUSION The potential mecha⁃nism of action of ICA against myocardial isch⁃emia-reperfusion injury may be related to the reg⁃ulation of processes such as CMEC migration and angiogenesis,and it functions through the key target gene PKC.

Protective effects of icariin on human umbilical vein endothelial cell injured by angiotensin Ⅱ

To investigate the effects of icariin （ICA） on angiotensin Ⅱ（Ang Ⅱ）-induced injury in human umbilical vein endothelial cells line （ECV-304）. The ECV-304 cells were cultured in vitro. After 24 h incubating with icariin, the model of AngⅡ-induced injury in ECV-304 was established. The cell viability （MTT method）, Lactate dehydrogenase （LDH） release and Nitric oxide （NO） production in the medium, the capacity of scavenging superoxide anion radicals （O2^-） and hydroxyl radicals （.OH） were measured. The activities of superoxide dismutase （SOD）, total nitric oxide synthase （T-NOS）, inducible nitric oxide synthase （iNOS） and constitutive nitric oxide synthase （cNOS） in the cells were determined. Compared with the Ang Ⅱ-treated group, ICA can significantly raise the viability of EC, increase the activities of SOD, T-NOS and cNOS, increase the production of NO, enhance the capacity of scavenging superoxide anion radicals （ O2^- ） and hydroxyl radicals（.OH）, and lower LDH leakage and iNOS activity. The results suggest that ICA can protect endothelial cells （ECV-304） from Ang II-induced injury.

Preparation and characterization of icariin/PHBV drug delivery coatings on anodic oxidized titanium

A composite material was fabricated by applying a biodegradable drug delivery coating,consisting of poly（3-hydroxyburyrate-co-3-hydroxyvalerate）（PHBV） and icariin,to an anodic oxidized titanium plate.The coating was prepared by evaporating chloroform solution containing PHBV and icariin on the titanium plate under vacuum condition.Icariin/PHBV coated titanium plates significantly enhance the proliferation of MG-63 cells compared with the PHBV coated and anodic oxidized ones.Increased icariin contained in the coating displays an elevated influence on cell proliferation.The results show that icariin gradually releases from the coating to cells mainly through the phospholipid-based cellular membrane instead of the culture medium.The overall results suggest that the novel icariin/PHBV coating can be used to enhance the bioactivity of titanium based orthopedic implants.

Icariin/HAP纳米凝胶剂的处方优化及体外释放研究

[目的]对Icariin/HAP纳米凝胶剂处方进行优化并考察其体外释放特性。[方法]物理吸附法制备Icariin/HAP纳米混悬液,研和法制备Icariin/HAP纳米凝胶,采用正交设计对处方进行优化,透析法测定Icariin/HAP纳米凝胶中药物的释放。[结果]Icariin/HAP纳米凝胶优选处方为卡波姆浓度为1%,三乙醇胺用量为1.5%,丙二醇用量为5%。Icariin/HAP纳米凝胶剂、Icariin凝胶剂和Icariin溶液体外释放均符合一级模型和Weibull模型。[结论]以卡波姆为基质制备Icariin/HAP纳米凝胶,处方简单,质量稳定,可提高Icariin/HAP纳米混悬液的稳定性。

Pharmacological effects of icariin and icariinriside on central nervous system

Epimedium Brevicornum is a traditional Chinese medicinal plant possessing properties of sweet, warm, tonifying kidney, strong bones and rheumatism. Icariin, a flavonoid compound, is one of the main active ingredients of Epimedium. Icariinriside(ICS) is the main metabolite of icariin. Icariinand ICS have multiple pharmacological effects such as anti-tumor, anti-oxidative stress, improvement of cardiovascular and cerebrovascular, and regulation of endocrine. We have conducted a series of studies on the neuroprotection and mechanisms of action of icariin and ICS for many years. The main findings are reported as follows.(1) Effect on Alzheimer disease(AD) model animals: Icariin significantly attenuated learning and memory loss, hippocampal neuron loss and senile plaque formation in APP/PS1 transgenic AD model mice, which may be related to inhibition of Aβ production and reduction of PDE5(phosphodiesterase 5).In addition, icariin significantly attenuated Aβ25-35-induced learning and memory decline and hippocampal neuronal apoptosis in rats, which may be related to lowering PDE5 content and up-regulating BDNF/Trkb/CREB signaling pathway, inhibiting MAPK and NF-κB signaling pathways, and increasing expression of acetylcholinesterase(ACHE) and choline acetyltransferase(CHAT) in the hippocampus. At the same time, icariin can significantly improve the learning and memory dysfunction induced by amanita proline in rats, which may be related to the inhibition of hippocampal neuronal apoptosis, antiexcitatory amino acid toxicity and regulation of MAPK and NF-κB signaling pathways.(2) Effects on Parkinson disease(PD) model animals: The study found that in LPS-induced dopaminergic neuron injury animal models and cell models, icariin can inhibit microglia by inhibiting the expression of inflammatory factors such as TNF-α, IL-1β, NO and COX-2. Activation of glial cells increases the expression of neurotrophic factors such as BDNF and GDNF, increases the content of dopamine(DA) and its metabolites 3, 4-dihydroxyphenylacetic acid(DOPAC) and homovanillic acid(HVA), inhibits MAPK and the NF-κB signaling pathway, protecting dopaminergic neurons. In addition, icariin significantly attenuated6-OHDA-induced dopaminergic neuronal damage. In Nrf2 knockout mice, the neuroprotective effect of icariin disappeared, suggesting that Nrf2 may be one of the targets of icariin to play neuroprotective effects.(3) Effects on vascular dementia(VD) model animals: Icarin can improve the learning and memory ability and memory function of chronic hypoperfusion rats, and its mechanism may be related to increase the level of VEGF/VEGFR2 protein in the brain and activate multiple downstream signaling pathways to promote angiogenesis to play an indirect protective effect on neurons;The level of BDNF/Trk B protein in the brain increases the phosphorylation level of CREB and exerts direct neuroprotective effects.(4)Effect on cerebral ischemia: In a model of ischemic brain injury, icariin acts to up-regulate Sirt1 by activating p38, thereby exerting an anti-ischemic injury and protecting neuronal cells. In addition, icariin has neuroprotective effects on cerebral ischemia-reperfusion injury in rats, which may increase GSH-Px,SOD activity, decrease MDA content, inhibit free radical damage, reduce NO content, NOS activity,and inhibit neurotoxic damage. Reduction of MPO activity, TNF-α, IL-1β content is associated with inhibition of inflammatory response.(5) Cell protection: Icariin has a protective effect on 6-OHDA-induced oxidative damage in PC12 cells, which may be related to inhibition of apoptosis and regulation of Keap1/Nrf2/ARE signaling pathway, while ICS can attenuate oxygen-glucose deprivation/reoxygenation-induced cellular damage in PC12 cells. The mechanism of cellular oxidative damage may be related to inhibition of apoptosis and regulation of Nrf2/SIRT3 signaling pathway.Icariin and ICS have good preventive and therapeutic effects on central nervous system diseases such as AD, PD, VD, etc. However, due to the complexity of the molecular mechanisms of icariin and ICS, the molecular mechanisms of the central nervous system are still worthy of further study.

Effects of icariin on cGMP-specific PDE5 and cAMP-specific PDE4 activities

Aim: To clarify the mechanism of the therapeutic action of icariin on erectlile dysfunction (ED). Methods: PDE5 was isolated from the human platelet and PDE4 from the rat liver tissue using the FPLC system (Pharmacia, Milton Keynes, UK) and the Mono Q column. The inhibitory effects of icariin on PDE5 and PDE4 activities were investigated by the two-step radioisotope procedure with [3H]-cGMP/[3H]-cAMP. Papaverine served as the control drug. Results: Icariin and papaverine showed dose-dependent inhibitory effects on PDE5 and PDE4 activities. The IC50 of Icariin and papaverine on PDE5 were 0.432 μmol/L and 0.680μmol/L, respectively and those on PDE4, 73.50 μmol/L and 3.07μmol/L, respectively. The potencies of selectivity of icariin and papaverine on PDE5 (PDE4/PDE5 of IC50) were 167.67 times and 4.54 times, respectively. Conclusion: Icariin is a cGMP-specific PDE5 inhibitor that may be developed into an oral effective agent for the treatment of ED.

Effects of icariin on erectile function and expression of nitric oxide synthase isoforms in castrated rats

Aim： To investigate the effect of icariin on erectile function and the expression of nitric oxide synthase （NOS） isoforms in castrated rats. Methods： Thirty-two adult male Wistar rats were randomly divided into one shamoperated group （A） and three castrated groups （B, C and D）. One week after surgery, rats were treated with normal saline （groups A and B） or oral icariin （1 mg/[kg·day] for group C and 5 mg/[kg·day] for group D） for 4 weeks. One week after treatment, the erectile function of the rats was assessed by measuring intracavernosal pressure （ICP） during electrostimulation of the cavernosal nerve. The serum testosterone （ST） levels, the percent of smooth muscle （PSM） in trabecular tissue, and the expression of mRNA and proteins of neuronal nitric oxide synthase （nNOS）, inducible nitric oxide synthase （iNOS）, endothelial nitric oxide synthase （eNOS） and phosphodiesterase V （PDES） in corpus cavernosum （CC） were also evaluated. Results： ICP, PSM, ST and the expression of nNOS, iNOS, eNOS and PDE5 were significantly decreased in group B compared with those in group A （P 〈 0.01）. However, ICE PSM and the expression of nNOS and iNOS were increased in groups C and D compared with those in group B （P 〈 0.05）. Changes in ST and the expression of eNOS and PDE5 were not significant （P 〉 0.05） in groups C and D compared with those in group B. Conclusion： Oral treatment with icariin （〉 98.6 % purity） for 4 weeks potentially improves erectile function. This effect is correlated with an increase in PSM and the expression of certain NOS in the CC of castrated rats. These results suggest that icariin may have a therapeutic effect on erectile dysfunction.

The testosterone mimetic properties of icariin

Aim： To evaluate the testosterone mimetic properties of icariin. Methods： Forty-eight healthy male Sprague-Dawley rats at the age of 15 months were randomly divided into four groups with 12 rats each： the control group （C）, the model group （M）, the icariin group （ICA） and the testosterone group （T）. The reproductive system was damaged by cyclophosphamide （intraperitoneal injection, 20 mg/kg·day） for 5 consecutive days for groups M, ICA and T, at the sixth day, ICA （gastric gavage, 200 mg/kg·day） for the ICA group and sterandryl （subcutaneous injection, 5 rag/rat.day） for the T group for 7 consecutive days, respectively. The levels of serum testosterone, luteinizing hormone （LH）, follicle stimulating hormone （FSH）, serum bone Gla-protein （BGP） and tartrate-resistant acid phosphatase activity in serum （StrACP） were determined. The histological changes of the testis and the penis were observed by microscope with hematoxylin-eosin （HE） staining and terminal deoxynucleotidyl transferase biotin-dUTP-X nick end labeling （TUNEL）, respectively. Results： （1） Icariin improved the condition of reproductive organs and increased the circulating levels of testosterone. （2） Icariin treatment also improved the steady-state serum BGP and might have promoted bone formation. At the same time, it decreased the serum levels of StrACP and might have reduced the bone resorption. （3） Icarrin suppressed the extent of apoptosis of penile cavernosal smooth muscle cells. Conclusion： Icariin has testosterone mimetic properties and has therapeutic potential in the management of hypoandrogenism.

Compound of icariin,astragalus,and puerarin mitigates iron overload in the cerebral cortex of Alzheimer＇s disease mice

Increasing evidence indicates that disruption of normal iron homeostasis may contribute to pathological development of Alzheimer＇s disease.Icariin,astragalus,and puerarin have been shown to suppress iron overload in the cerebral cortex and improve spatial learning and memory disorders in Alzheimer＇s disease mice,although the underlying mechanism remains unclear.In the present study,APPswe/PS1ΔE9 transgenic mice were administered icariin,astragalus,and puerarin（120,80,and 80 mg/kg,respectively,once a day,for 3 months）.Iron levels were detected by flame atomic absorption spectroscopy.Interleukin-1β,interleukin-6,and tumor necrosis factor-α levels were measured in the cerebral cortex by enzyme linked immunosorbent assay.Glutathione peroxidase and superoxide dismutase activity and malondialdehyde content were determined by colorimetry.Our results demonstrate that after treatment,iron levels and malondialdehyde content are decreased,while glutathione peroxidase and superoxide dismutase activities are increased.Further,interleukin-1β,interleukin-6,and tumor necrosis factor-α levels were reduced.These results confirm that compounds of icariin,astragalus,and puerarin may alleviate iron overload by reducing oxidative stress and the inflammatory response.

Relationships Between Icariin and Anti-Apoptotic miRNA-21 in Mouse Blastocyst Development In vitro

In this study, the effect of icariin, a flavonoid from the Chinese traditional medicine epimedium, on miRNA-21 of mouse developmental blastocysts in vitro and the development of preimplantation embryos were studied. The possible effective targets of icariin promoting preimplantation embryo development in vitro and anti-apoptosis were determined. The embryos were cultured in modified CZB medium （mCZB） as control group. The experimental group （Ica group） was supplemented with 0.6 μg mL-1 icariin. Mouse pronuclear embryos were cultured in vitro until blastocysts. The development rates of preimplantation embryos were observed. The total cell number, apoptotic cell number and the rate of apoptotic cells in blastocysts were analysed by the staining of Hoechst33342 and labeling of TUNEL and detected under a laser confocal scanning microscope. The miRNA-21 expression, the mRNA levels of pro-apoptotic Caspase3, and the target genes of miRNA-21： pro-apoptotic PTEN, anti-apoptotic Bcl-2 were detected by real-time RT-PCR. The results showed that percentages of morulaes and blastocysts in Ica group were both extremely higher than control group （（85.14±6.57）% vs. （72.04±11.58）%; （82.50±7.11）% vs. （66.80±11.70）%, respectively, P〈0.01）. The total cell number ofblastocysts had extreme difference between Ica group and control group （（61.40±9.64） vs. （46.23±4.50）, P〈0.01）. The apoptotic cell number and rate of apoptotic cells of blastocysts were both reduced in Ica group （（1.47±0.51） vs. （2.94±0.66）; （2.40±0.27）% vs. （6.25±0.62）%, respectively, P〈0.01）. Compared to control group, addition of icariin into mCZB extremely increased the expression of anti-apoptotic miRNA-21 （P〈0.01）, down-regulated pro-apoptotic Caspase3 （P〈0.05） and PTEN （P〈0.01）, up-regulated anti-apoptotic Bcl-2 （P〈0.01）. In conclusion, icariin could reduce the apoptosis, promote the embryo development in vitro by enhancing miRNA-21 expression to up-regulated anti-apoptotic genes and down-regulated pro- apoptotic genes. These apoptosis-related genes were regulated by miRNA-21.

Inhibitory effect of icariin on expression of myelin inhibitory factors in the central nervous system of rats with focal cerebral ischemia

Icariin, the major active component of Chinese medicinal herb epimedium brevicornum maxim, is used widely in traditional Chinese medicine for the treatment of neurological diseases. However, the effects of icariin on myelin inhibitory factors are as yet unclear. In the present study, administration of icariin at 20 mg/kg showed a marked reduction in neurological deficit of middle cerebral artery occlusion rats. Icariin exhibited better inhibitory effects on myelin inhibitory factors： Nogo-A, myelin-associated glycoprotein and oligodendrocyte myelin glycoprotein in ischemia regions of middle cerebral artery occlusion rats compared with monosialotetrahexosylganglioside. These results indicate that icariin exhibits potent inhibitory effects on expression of myelin inhibitors after middle cerebral artery occlusion-induced focal cerebral ischemia in vivo. This effect may be mediated, at least in part, by the inhibition of both Nogo-A, myelin-associated glycopretein and oligodendrocyte myelin glycoprotein activation, followed by the enhancement of axonal sprouting and regeneration, resulting in neurological functional recovery.